Stabilisation of Recombinant Aequorin by Polyols: Activity, Thermostability and Limited Proteolysis

The photoprotein aequorin is a calcium-dependent bioluminescent enzyme which is most widely used in biotechnology processes, but this protein is susceptible to aggregation and proteolysis degradation. Various additives such as polyols are known to enhance the stability of proteins and protect them in native folded and functional state. In this work, for study of aequorin stability, the histidine-tagged apoaequorin was expressed in Escherichia coli and purified by nickel chelate affinity chromatography. Kinetics of light emission of purified aequorin upon addition of Ca²⁺ showed a linear dependency on aequorin concentration. Furthermore, the effect of some stabilisers, such as glycerol, glucose, lactose, terehalose, sucrose and sorbitol on thermostability of recombinant aequorin was measured. Results indicate that the recombinant aequorin is very stable in phosphate buffer including 30 mM sorbitol, since after heat shock of 30 min at different temperatures, a slight decrease in activity was observed. However, flexibility and exposure of tryptophan residues of aequorin to the solvent, in the presence and absence of stabilisers, with respect to fluorescence quenching by acrylamide, indicated identical characterisation. In addition, according to limited proteolysis of aequorin demonstrating that this enzyme is sensitive to proteases as in the presence of 2 ng/ml of protease, aequorin was completely digested. In conclusion, sorbitol increases stability of aequorin with high photoactivity and not effect for flexibility and limited proteolysis of this photoprotein.     

Comparative thermostability of whey protein and alginate hydrospheres complexed with divalent cations

Journal of Thermal Analysis and Calorimetry - Water retention and thermostability of powders and films of whey protein isolate (WPIp, dWPIf), sodium alginate (NaALGp, NaALGf), and their dry...     

联苯并环丁烯的制备及其聚合物的热稳定性

苯并环丁烯树脂具有优异的综合性能,可作为微电子器件中的高性能介电薄膜材料。其优异的性能满足大规模集成电路(MCM)、微电机系统(MEMS)、液晶显示器等器件的苛刻性能要求,在微电子领域有着极为重要的应用且前景广阔[1,2,3]。苯并环丁烯单体的种类较多,其聚合物大多具有优异的     

脱辅基神经红蛋白热稳定性的光谱法研究

采用酸丙酮法制备了脱辅基神经红蛋白(apo-Ngb)。利用紫外-可见吸收光谱和圆二色光谱(CD)对apo-Ngb的热稳定性进行了研究。结果表明,在高温下apo-Ngb发生了变性,其二级结构遭到破坏,α-螺旋的含量减少。计算得到了apo-Ngb热变性的中点温度(61.8℃)和热力学参数(ΔH=93.1 kJ·mol-1,ΔS=277.8J·mol-1·K-1)。与Ngb相比,由于血红素辅基的去除,apo-Ngb蛋白肽链结构变得疏松无序,从而使apo-Ngb的热稳定性有所降低。     

Phanerochaete mutants with enhanced ligninolytic activity

In addition to lignin, the white rot fungusPhanerochaete chrysosporium has the ability to degrade a wide spectrum of recalcitrant organopollutants in soils and aqueous media. Most of the organic compounds are degraded under ligninolytic conditions with the involvement of the extracellular enzymes, lignin peroxidases, and manganese-dependent peroxidases, which are produced as secondary metabolites triggered by conditions of nutrient starvation (e.g., nitrogen limitation). The fungus and its enzymes can thus provide alternative technologies for bioremediation, biopulping, biobleaching, and other industrial applications. The efficiency and effectiveness of the fungus can be enhanced by increasing production and secretion of the important enzymes in large quantities and as primary metabolites under enriched conditions. One way this can be achieved is through isolation of mutants that are deregulated, or are hyperproducers or supersecretors of key enzymes under enriched conditions. Through UV-light and γ-ray mutagenesis, we have isolated a variety of mutants, some of which produce key enzymes of the ligninolytic system under high-nitrogen growth conditions. One of the mutants, 76UV, produced 272 U of lignin peroxidases enzyme activity/L after 9 d under high nitrogen (although the parent strain does not produce this enzyme under these conditions). The mutant and the parent strains produced up to 54 and 62 U/L, respectively, of the enzyme activity under lownitrogen growth conditions during this period. In some experiments, the mutant showed 281 U/L of enzyme activity under high nitrogen after 17 d.

     

Fluorescence studies of protein thermostability in ionic liquids

Using the single tryptophan residue in the sweet protein monellin as a spectroscopic handle, we show the extreme thermodynamic stabilization offered by an ionic liquid; T(un) approximately 105 degrees C in [C4mpy][Tf2N] compared to 40 degrees C in bulk water.     

Novel teststrip with increased accuracy

A new type of a simple and cheap teststrip for liquid samples is described. It is based on microchromatography or microtitration on a porous, capillary-active substrate (e.g., filter paper or a similar absorbent material). In case of microchromatography an analyte-selective indicator (and other auxiliary reagents) is impregnated or immobilized on the capillary-active substrate; in case of micro-titration the capillary-active substrate contains a titrant which reacts stoichiometrically with the analyte. Quantitative analysis is performed by measurement of an area (which had changed its color) rather than by evaluation of the shade or intensity of a color (like in conventional teststrips). In order to show the broad applicability of this new principle teststrips for different analytes like Quaternary Ammonium Compounds (QACs), the cation Ni2+, the anion SO4(2-) and H2O2 are described. The detection limit and working range of the novel teststrip can be adjusted by variation of its size.     

Increased thermostability of non-polar wall-coated open tubular columns

A method of deactivating the inner surface of glass capillary columns is described, which can be used at 350°C without detoriation. A non-polar liquid phase was prepared from a commercially available liquid phase which, when coated on a glass surface, can withstand temperatures of 325°C for isothermal analysis and 350°C for temperature-programmed analysis. After deactivation, the column was coated using the static coating method. Then it was conditioned, tested and kept for 48 hours at 35O°C before being used for the analysis of a mixture of chlorinated pesticides.     

Synthesis and thermostability of the binary condensed phosphates

Some condensed phosphates, especially cyclo-tetraphosphates have been synthetized as new binary compounds and their existence has been proved. The synthesis of cyclo-CdMgP₄O₁₂ is shown as an example. The products crystallize in the monoclinic system (C2c group) and are stable to 900°C. The synthesis is based on a thermal procedure making use of the reversible transformation of cyclo-tetraphosphates to higher linear phosphates (the 1.method) and on a calcination of the starting binary dihydrogenphosphate at quasi-isothermal-isobaric conditions (the 2.method).

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Zusammenfassung Einige kondensierte Phosphate, im besonderen Zyklotetraphosphate wurden als neue binäre Verbindungen hergestellt. Als Beispiel wird Zyklo-Cd MgP₄ O₁₂ angeführt. Das Produkt kristallisiert monoklin /Gruppe C2/c/ und ist bis 900°C stabil. Die Synthese basiert auf einem thermischen Verfahren unter Anwendung des reversiblen Überganges von Zyklotetraphosphaten zu höhereren linearen Phosphaten /Verfahren 1/ und auf dem Kalzinieren der binären Ausgangsverbindung Dihydrogenphos phat bei quasi-isothermen-isobaren Bedingungen /Verfahren 2/.

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$ , . - dMgP₄O_I2. $ / 2/ $ 900°. $ $ , — $ - .

     

Correction to: Comparative thermostability of whey protein and alginate hydrospheres complexed with divalent cations

Journal of Thermal Analysis and Calorimetry -     

The first systhesis of a chromia-pillared layered niobate with porosity, acidity and high thermostability

Although the hydrated niobic acid is an unusual solid acid and making an impact onits application in catalysis, it is usually nonporous and has a rather low surface area andweak acidity upon heat treatment. To overcome these shortcomings, the layered nio-bate had been pillared with thermostable inorganic oxides such as alumina and silica.Chromia is also a suitable inorganic oxide which can be used as pillar and contributesnew properties to the resultant materials. Here, we report the first synthesis of achromia-pillared layered niobate with porosity, acidity and thermostability.     

Structural adhesives with increased deformation resistance

The radiation-resistant cold-setting adhesive compositions with increased elasticity have been developed based on epoxy-diane resin modified by low-molecular two- and three-functional diluents. The presented adhesives can operate in the temperature range of −196 to +150°C, and they yield a high shear strength of the adhesive joint for aluminum alloys (16–23 MPa) and peeling strength for the joint between organic fabrics and aluminum alloys (1.75–2.4 kN/m).     

Metal-organogermanate frameworks built by two kinds of infinite Ge-O chains with high thermostability and luminescent properties

A bifunctional metalloligand with metal-C (Ge-C) bonds, bis(carboxyethylgermanium) sesquioxide (H(2)E(2)Ge(2)O(3), where E = -CH(2)CH(2)COO(-)), has been used to make metal-organometallic frameworks (MOMFs) Cd(1.5)(E(3)Ge(3)O(5)) (1) and Pb(E(2)Ge(2)O(3)) (2). In the structures, the inorganic Ge(2)O(3) cores of the metalloligands polymerize to form two kinds of infinite Ge-O chains, and the organic carboxyls induce the Cd and Pb ions to produce Cd-O and Pb-O chains. Further, the Ge-O chains link the Cd-O/Pb-O chains via -CH(2)CH(2)- groups to result in two novel MOMFs, showing high thermostability and blue-violet emission.     

The influence of polyvalent cations on the thermostability of cellulose

The influence of some polyvalent cations on the thermostability of cellulose was investigated using both conventional TG analysis and quasi-isothermal treatment at elevated temperatures. It was established that ions of copper, cobalt and iron absorbed in the cellulose fibre surface accelerated the thermal decomposition of cellulose chain molecules, the temperature of the main decomposition decreasing very significantly. Lewis acid type additives (such as ZnCl₂) promote the hydrolytic decomposition and hence the temperature of the post-decomposition increases compared with that of the blank sample. Ions of alkaline earth metals (Ca and Ba) do not affect the thermal behaviour of cellulose fibres. From the results of the quasi-isothermal experiments the apparent energy of activation for the decomposition of cellulose in the presence of metal ions was calculated.     

Synthesis of volatile alkoxygallium hydrides and study of their thermostability

A method for the synthesis of alkoxygallium hydrides (ROGaH₂ and (RO)₂GaH, where R=Prⁱ, Buⁱ) was proposed. The method is based on the reaction of gallane GaH₃ with one or two equivalents of the corresponding alcohol. Thermolysis of these compounds was studied by differential thermogravimetry. Translated fromIzvestiya Akademii Nauk. Seriya Khimicheskaya, No. 8, pp. 1655–1657, August, 1998.     

Amphiphilic copolymers with increased affinity for substrates

The copolymers of methyl quaternized 2-dimethylaminoethyl acrylate and styrene, 2-vinyl naphthalene, acrylic acid iso-octyl ester, or acrylic acid n-butyl ester have been prepared. Studies were made of the binding of a “binding probe,” methyl orange, by the copolymers in aqueous solution. The first binding constants and the thermodynamic parameters in the course of the binding were evaluated. Furthermore, the intensity of fluorescence of a hydrophobic fluorescent probe, 2-p-toluidinylnaphthalene-6-sulfonate, in the presence of these polymers was investigated. In addition, the fluorescence spectra of monomer and excimer emissions of the polymers with aromatic residues were measured. The excimer/monomer fluorescence intensity ratio was studied in the presence of various additives such as methyl orange, urea, methanol, and NaCl to gain an insight into the nature of microdomains in the polymer. The nature and phenomena of dye binding and hydrophobic fluorescent probe binding with the polymers are discussed. © 1993 John Wiley & Sons, Inc.     

Effect of polyvinylalcohols on the thermostability of lipase fromCandida rugosa

Lipase fromCandida rugosa was stabilized against thermal inactivation in the presence of polyvinylalcohols (PVA) of different molecular weights. The apparent rate constant of the lipase inactivation, k_d, at 49°C is 0.049/min and 0.022/min in the absence and in the presence of PVA (mol wt 22,000), respectively. The improvement of the lipase thermostability by adding PVA was confirmed by differential scanning calorimetry. The presence of PVA had also an effect on the hydrolytic activity of the enzyme. Furthermore, lipase was modified by covalent linkage to PVA by means of an original procedure. With respect to the native enzyme, the modified lipase has a slightly lower specific activity, but it is more stable against heat denaturation (k _d 0.032/min at 49°C).